The invention is related to the field of treatment of cancer, other neoplastic disorders, and other conditions in vertebrates in which killing a specific group of cells is useful.
Most if not all cells of metazoan animals carry the machinery to commit suicide in a regular manner in response to suitable stimulus. This process is called programmed cell death, cell suicide, or apoptosis. Apoptosis is being extensively studied in mammals and other vertebrates, as well as in the worm Caenorhabdits elegans and the fly Drosophila melanogaster (reviews: Ellis et al., 1991; Steller, 1995). In vertebrate cells the process of apoptosis, which was previously termed “shrinkage necrosis,” involves a regular sequence of events, including membrane blebbing, cell shrinkage, pycnosis of nuclei with margination of chromatin, and usually cleavage of DNA into nucleosome-sized fragments (Wyllie et al., 1980).
Apoptosis is an essential part of embryonic development and of the maintenance of an adult animal. In mammals, for example, during development apoptosis plays a major role in the development of the nervous system (more than 50% of the neural cells that arise during embryogenesis undergo apoptosis), in the elimination of lymphocytes that produce antibodies which recognize self, in “carving” features such as the digits of the hand, and so forth. Throughout life, orderly apoptosis is used to eliminate damaged or unwanted cells without inducing an inflammatory reaction. Blood cells, cells of the immune system, and cells of most if not all tissues normally are eliminated by the apoptotic mechanism.
Failures of apoptosis produce or contribute to severe diseases, including autoimmune diseases and some cancers. It has been argued that one of the major causes of the development and progression of many cancers is a reduction of the occurrence of apoptosis (Wyllie, 1985; Fisher, 1994; Hickman et al., 1994; Martin and Green, 1995; Thompson, 1995).
A wide variety of signals induce apoptosis in suitable target cells (Gerschenson and Rothello, 1992; Thompson, 1995). Radiation and many valuable chemotherapeutic agents, such as cisplatin and other platinum compounds, induce apoptosis (e.g., Eastman, 1990; Hickman, 1992; Chu, 1994a). These agents affect many cell types. Specialized cell types are dependent on specific growth factors (e.g., nerve growth factor for certain neuronal cells, interleukin-2 for certain lymphocytes) and undergo apoptosis if the required factors are unavailable. Other cell types have receptors for specific agents that can induce apoptosis in these cell types (e.g., glucocorticoid for thymocytes, tumor necrosis factor in suitable target cells) (e.g., Rubin et al., 1988).
The mechanism of apoptosis is just beginning to be understood. Some have suggested that all cells are poised to die, and that they are kept alive by constant “survival signals” that keep the suicide machinery inactive (Raff, 1992). It is clear that many if not all vertebrate cells contain preformed machinery for apoptosis, since there are many examples of cells that undergo apoptosis even without synthesis of new proteins (Waring, 1990). There also are cases in which protein synthesis is required (reviewed by Cohen, 1993).
Several elements that appear to be part of the apoptotic machinery have been identified and are receiving much attention. Two that should be mentioned are bcl-2 and its family members and p53. Exactly how these are related to the apoptotic machinery is still being defined.
Expression of oncogene bcl-2 in cells markedly delays or blocks induction of apoptosis by many agents, including some that are valuable in chemotherapy of tumors, such as cisplatin (Reed, 1994; Korsmeyer, 1995; Thompson, 1995). There are a few cases in which induction of apoptosis is unaffected by expression of bcl-2 (e.g., Sentman et al., 1991; Vaux et al., 1992). High-level expression of bcl-2 is common in tumors, including breast carcinomas, small cell lung cancer, androgen-independent prostate cancer, and neuroblastoma (Hickman et al., 1994). In some cases expression of bcl-2 is correlated with a poor prognosis for therapy (Reed, 1994).
Functional tumor suppressor gene product p53 is required for induction of cell death by irradiation and many chemotherapeutic agents (Lowe et al., 1993), as well as by oxygen deficiency at the center of solid tumors (Graeber et al., 1996). On the other hand, the normal development of transgenic animals nullizygous for the p53 gene indicates that p53 is not required for the extensive apoptosis that occurs during development (Donehower et al., 1992). Other cases of p53-independent apoptosis have been described (White, 1993; Zhuang et al., 1995). Many established lines of cells in culture have lost p53 function. In tumors in vivo, loss of p53 function is common, and this loss is correlated with tumor aggressiveness and indicates a poor prognosis for treatment by standard protocols of chemotherapy and radiation (Fisher, 1994; Hartmann et al., 1997).
As an example, the roles of p53 loss and bcl-2 expression in the development and progression of colon carcinomas have been described and analyzed (Hickman et al., 1994; Sinicrope et al., 1996).